Results
All birds had enlarged congestive greenish-black tinged livers with a reticulate pattern, enlarged kidneys and mottled locally pale pectoral muscles (Figure 1A-C). The distribution, type and severity of lesions varied between animals. In all cases, the most prominent microscopic lesions were hyperemia and congestion, necrosis and mononuclear inflammatory infiltrates consisting of lymphoid cells, plasma cells and histiocytes, although to a different degree depending on the tissue. The liver showed congested central veins, an enlarged sinusoidal area, hemozoin pigment within hyperplastic Kupffer cells, and dense lymphoplasmacytic and histiocytic inflammatory cells within the portal tract and in perivascular spaces (Figure 1D). However, the most striking finding in the liver was the great number of phanerozoites inside hepatic cells (Figure 1D, inset). Other organs affected included the heart and, especially, the kidney, where multifocal necrosis along with mononuclear and polymorphonuclear infiltrates were the most important lesions (Figure 1E). Phanerozoites were also observed in other organs, especially in the skin and muscle where they were associated to multifocal necrosis as well as mononuclear and polymorphonuclear infiltrates (Figure 1F). In the spleen, lymphoid depletion, necrotic foci of lymphoid cells, multifocal granulocytic infiltrates and a severe hemosiderosis were evident. In the lung, oedema and congestion, as well as moderate to severe diffuse interstitial bronchopneumonia were observed. Lesions in the central nervous system were characterized by oedema, congestion, gliosis, neuronal necrosis, perivascular cuffing, capillary endothelial cell swelling, and perivascular infiltrates composed of mononuclear cells.
Using immunohistochemistry, BAGV antigen was detected in numerous organs, including spleen, liver, lung, kidney, central nervous system and muscle (Figure 1G, H). Specifically, antigen was detected in the cytoplasm of neurons and glial cells, endothelial and epithelial cells, macrophages and hepatocytes, often associated to necrotic foci (Figure 1H). Plamodium spp. phanerozoites observed in the hematoxilin-eosin stained tissues were confirmed to be Plasmodium spp. by immunohistochemistry (Figure 1I).
Bagaza virus was identified by real time RT-PCR and subsequent sequencing in vascular feather and brain samples of the partridges (Figure 2), while all tested tissues (feather, brain, spleen, heart, liver) also tested positive for Plasmodium spp.